Peptide drugs are usually administered systemically, e.g. parenterally due to their poor oral absorption and high instability in gastric fluids. However, parenteral administration may be painful and cause discomfort, especially for repeated daily administrations. In order to minimize the number of injections to a patient, the drug substance is advantageously administered as a depot formulation. The parenteral administration of peptide drugs as a depot formulation in a biodegradable polymer, e.g. as microspheres or implants, has been proposed enabling their sustained release after a residence time in the polymer which protects the peptide against enzymatic and hydrolytic influences of the biological media. A common drawback with injectable depot formulations is the fluctuation in plasma levels such as high peak levels which cause undesired pharmacological side reactions together with plasma levels close to zero during the entire release period. A therapeutically relevant blood level over an extended period of time is difficult to achieve and satisfactory peptide release profiles are in practice only obtained in very few cases.
Identified factors that control the peptide release characteristics of a parenteral depot in the form of PLGA microspheres include peptide form (i.e., free peptide, salt form), polymer type (i.e., molecular weight, lactide to glycolide ratio, linear or branched structure, end-terminal groups), drug loading, particle size and particle porosity and the distribution of the drug into the polymer matrix (U.S. Pat. No. 5,538,739).
Few commercial long-acting release drug formulations of microspheres are available on the market. SANDOSTATIN LAR® is a commercial available parenteral depot formulation comprising octreotide acetate active peptide. It is indicated for, inter alia, long-term maintenance therapy in acromegalic patients, and treatment of severe diarrhea and flushing associated with malignant carcinoid tumors and vasoactive intestinal peptide tumors (vipoma tumors). Approved at doses of 10, 20 and 30 mg (and up to 40 mg for patients with acromegaly in certain countries such as the US and Japan), Sandostatin LAR® allows for once-monthly intragluteal injection. The pharmacokinetic profile of octreotide after injection of Sandostatin LAR® reflects the release profile from the polymer matrix and its biodegradation. After a single i.m. injection in humans, the octreotide concentration reaches a transient initial peak within 1 hour after administration, followed by a progressive decrease to a low undetectable level within 24 hours. After this initial release on day 1, octreotide remains at sub-therapeutic levels in the majority of the patients for the following 7 days. Thereafter, octreotide concentration increases again and around day 14 (about 2-3 weeks post-injection) reaches plateau levels which are maintained during the following 3 to 4 weeks, then a decline period of 6 weeks follows (Summary of product characteristics for Sandostatin LAR® 10 mg, 20 mg or 30 mg powder and solvent for suspension for intramuscular injection, 2013). In agreement with the above and according to the available literature data the expected release profile for Sandostatin LAR in rats follows the same pattern (AAPS PharmSciTech, Vol. 12, No 4 (2011)).
There are a number of techniques for the microencapsulation of peptides in PLGA microspheres. The most widely used techniques both in lab scale and for commercial productions include phase separation/coacervation technique, spray drying and single or double emulsion/solvent evaporation technique (PDA J Pharm Sci and Tech 2008, 62 125-154; Microencapsulation Methods and Industrial Applications Second Edition).                1. In phase-separation or coacervation technique, an aqueous solution of peptide/protein is emulsified or alternatively the peptide/protein is dispersed in solid form into solution containing dichloromethane and PLGA. Silicone oil is added to this dispersion at a defined rate, reducing solubility of polymer in its solvent. The polymer-rich liquid phase (coacervate) encapsulates the dispersed peptide/protein molecules, and embryonic microspheres are subjected to hardening and washing using heptane. The process is quite sensitive to polymer properties, and residual solvent is also an important issue.        2. In spray-drying technique a polymer is dissolved in a volatile organic solvent such as dichloromethane or acetone. The protein is suspended as solid or emulsified as aqueous solution in this organic solution by homogenisation. After that, the resulting dispersion is atomised through a (heated) nozzle into a heated air flow. The organic solvent evaporates, thereby forming microspheres with dimensions of typically 1-100 m. The microspheres are collected in a cyclone separator. For the complete removal of the organic solvent, a vacuum-drying or lyophilization step can follow downstream. The internal structure of the resulting polymeric microspheres depends on the solubility of the peptide/protein in the polymer before being spray-dried leading to the formation of reservoir or matrix type products. When the initial dispersion is solution, the final product obtained following spray drying is matrix or monolithic type, that is, polymer particles with dissolved or dispersed nature of the active ingredient (defined as microspheres). Conversely, when the initial dispersion is in suspension, the product obtained is reservoir type, that is, a distinct polymeric envelope/shell encapsulating a liquid core of dissolved active ingredient (defined as microcapsules).        3. Oil-in-water (o/w) and water-in-oil-water (w/o/w) are the two hydrous techniques representing, respectively the single and double emulsion formation during microspheres preparation. In these processes, peptides/proteins are dissolved in an organic solvent (e.g., alcohol) or in an aqueous solution and then mixed or emulsified with an organic solution (non-miscible with water) of the polymer to form a solution or water-in-oil (w/o) emulsion, respectively. Dichloromethane serves as organic solvent for the PLGA and the o/w primary emulsion is formed using either high-shear homogenization or ultrasonication. This primary emulsion is then rapidly transferred to an excess of aqueous medium containing a stabilizer, usually polyvinyl alcohol (PVA). Again homogenization or intensive stirring is necessary to initially form a double emulsion of w/o/w. Subsequent removal (by evaporation) of organic solvent by heat, vacuum, or both results in phase separation of polymer and core to produce microspheres. Instead of solvent evaporation, solvent extraction with large quantity of water with or without a stabilizer can also be undertaken to yield microspheres containing peptide/protein. Although the w/o/w microencapsulation technique seems to be conceptually simple to carry out, the particle formation process is quite complicated, and a host of process parameters are having an influence on or affect the properties of peptide/protein-loaded PLGA microspheres.        
Until now the temperature profile as applied during the evaporation step in emulsion/solvent evaporation techniques has not been identified as a critical process parameter to affect the release characteristics of the peptide from the polymer matrix. On the contrary, processing under constant temperature slightly above the boiling point of the organic solvent is generally applied (or slightly above the vapour pressure of the solvent when a reduced pressure/vacuum is applied to accelerate the evaporation of the solvent).
A typical release mechanism for these types of formulations includes three phases that can be generally represented as the initial release phase (phase 1), the hydration phase (phase 2), and primary release phase (phase 3) that is diffusion controlled but facilitated by erosion of the polymer matrix. The drug release begins after a lag time when the polymer molecular weight falls below a critical value and thus mass loss can take place (Faisant N, Siepmann J, Benoit J P. PLGA-based microspheres: elucidation of mechanisms and a new, simple mathematical model quantifying drug release. Eur J Pharm Sci. 2002 May; 15(4):355-66; Körber M. PLGA erosion: solubility- or diffusion-controlled? Pharm Res. 2010 November; 27(11):2414-20). There is a need in the art for an improved manufacturing method to control the release profile of the peptide drug substance from the polymer matrix of the microspheres.